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NEOPLASIA
From the Center for Cell and Gene Therapy, Baylor
College of Medicine, Houston, TX.
Because tumor-specific antigens have been identified in multiple
myeloma (MM), immunotherapy might provide an additional treatment modality for the disease. Expression of CD40 ligand (CD40L) proximate to the MM cells might serve this purpose, either by increasing their
capacity to present self-antigens by activation through their CD40
receptor or by the recruitment of professional antigen-presenting cells
(APCs) able to take up and present tumor-associated antigens. To
distinguish between these possibilities and predict whether human
CD40 High-dose chemotherapy followed by autologous stem
cell transplantation has improved the overall survival of multiple
myeloma (MM) patients.1,2 However, almost all patients
achieving complete remission ultimately experience
relapse,1,2 and immunotherapy may represent a means of
maintaining complete remission. Several antigens have been identified
as potential targets of T- and/or B-cell immune responses in MM,
including the idiotype of monoclonal immunoglobulin secreted by
malignant plasma cells,3-5 the underglycosylated form of
MUC-1,6 and antigens encoded by the MAGE-type
genes and by the FRAME gene.7
Most clinical studies of immunotherapy for MM have targeted the
specific idiotype.8-11 This specific tumor-associated
antigen has been used directly as a protein8 or DNA
vaccine9 or used to pulse dendritic cells in
vitro.10 Although this strategy can induce an
anti-idiotype immune response, little cytotoxic-effector function
against myeloma cells is recruited.8-11 Whole tumor cells, rather than single antigens, may be more likely to recruit a potent and
efficient cytotoxic antitumor response, because they may provide both
class I and class II major histocompatibility complex (MHC)-restricted antigenic epitopes, allowing interaction between CD4+ and
CD8+ T cells.12 To favor recruitment and
activation of immune effector cells, tumor cells have been genetically
modified to produce immunomodulatory molecules, such as
granulocyte-macrophage colony-stimulating factor or interleukin-2
(IL-2).13-15 Paracrine production of these molecules results in local recruitment of professional antigen-presenting cells
(APCs)16 that process tumor antigens derived from the irradiated tumor cells injected as the vaccine and help generate a
systemic antitumor immune response.13-16 Other
immunostimulatory molecules may be equally effective. CD40 ligand
(CD40L, CD154) is a member of the tumor necrosis factor gene family; it
is expressed mainly on antigen-activated CD4+ T lymphocytes
and plays a crucial role initiating antigen-specific cytotoxic T-cell
response.17 In addition, the interaction between CD40L and its receptor (CD40) expressed on the surface of professional APCs, induces these cells to mature, enabling them to directly recruit
and activate CD8+ T lymphocytes.17-20 For this
reason, transgenic expression of CD40L by tumor cells is currently
being explored as an antitumor vaccine.21-23
Transgenic expression of CD40L may also directly affect tumor cells
themselves. Tumor cells may fail to present antigens because they lack
the costimulatory molecules (such as B7-1/CD80 and B7-2/CD86) necessary
for activation of the T-cell receptor complex.24
Up-regulation of these costimulatory molecules on the surface of tumor
cells may therefore enhance their immunogenicity.24,25
Many B-cell tumors, including follicular lymphoma, lymphoblastic
leukemia, chronic lymphocytic leukemia (CLL), and some MMs, express the CD40 antigen on their surface; in lymphoma/CLL cells, the engagement of
CD40L has been shown to increase CD80/B7.1 and CD86/B7.2 expression, up-regulate HLA class II molecules and cell-adhesion molecules, and
enhance their antigen-presenting capability.21,26-28
Because CD40L may enhance antigen presentation both by tumor cells and
by professional APCs, we investigated whether transgenic CD40L
expression in a murine model of MM was protective. Because human MM may
be either CD40+ or CD40 Cell lines and cultures
Vectors and fibroblast transduction
Animal experiments In vivo experiments were performed in female BALB/CBYJ mice aged 8 to 10 weeks (The Jackson Laboratory, Bar Harbor, ME) syngeneic with all 3 plasmacytoma cell lines and with the CL7.1 fibroblast cell line. Animals were housed under specific pathogen-free conditions and treated according to the Baylor College of Medicine (Houston, TX) animal husbandry guidelines.In vivo tumor growth To assess the effect of CD40L transgene expression on tumor growth, mice were inoculated with 2 × 106 MPC-11 or S107 cells mixed with 1 × 106 CL7.1/mCD40L or CL7.1/neo fibroblasts (ratio, 2:1) on day 1. Fibroblasts were -irradiated
(1000 cGy) immediately before injection. Because unmodified MPC-11 and
S107 plasmacytoma cell lines are highly aggressive in
vivo,30,31 cells were sublethally -irradiated (3500 cGy) immediately before injection. The X-24 cell line has lower
tumorigenicity compared with MPC-11 and S107 cell lines and was
administered in correspondingly higher numbers (4 to
6 × 106 cells per animal) and without irradiation. The
tumor cell/fibroblast mixture was washed in phosphate-buffered saline
(PBS) (GIBCO BRL, Grand Island, NY), resuspended in 200 µL
final volume, and injected subcutaneously (SC) into the right leg.
Tumor growth was monitored 2 or 3 times a week by measuring the 2 maximum diameters of the tumor at the site of inoculation with a
caliper and was reported as a mean of the 2 diameters. In accordance
with the Animal Husbandry Committee Guidelines of the Animal Facility
at Baylor College of Medicine, the animals were killed when the tumor
size was larger than 10% of the animal's body weight or when the
tumor induced distress or ulceration.
In vivo T-lymphocyte depletion To evaluate the role of T lymphocytes in reducing tumor burden, mice were depleted of T lymphocytes by intraperitoneal (IP) inoculation of Gk1.5 (anti-CD4) and 2.43 (anti-CD8) monoclonal antibodies (mAbs) kindly provided by Dr Peter Doherty (St Jude Children's Research Hospital, Memphis, TN). The mAbs were inoculated IP with sublethally irradiated plasmacytoma cells mixed with CL7.1/mCD40L the day before the injection and then twice a week for 4 weeks. This protocol depletes more than 98% of the relevant T-cell subset in mice,21 an efficiency confirmed in these studies by FACS analysis of isolated splenocytes obtained from euthanized animals.Tumor-challenge studies Mice were inoculated SC with CL7.1/mCD40L fibroblasts mixed with sublethally irradiated plasmacytoma cell lines on the right leg on day 1 and in the opposite leg on day 14. At 2 weeks after the second injection, these animals were challenged on the back with the corresponding live plasmacytoma cell line. We used 5 × 105 cells per mouse for MPC-11 and S107 cell lines and 2 to 6 × 106 cells per mouse for the X-24 cell line.30,31 As a control, an equal number of live cells were injected in the backs of nonimmunized animals. Tumor growth was followed by measurement of the 2 maximum diameters of the tumor at the site of challenge, and mice were killed when the tumor size was larger than 10% of the animal's body weight or had induced distress or ulceration. To evaluate the specificity of the immune response, we also performed cross-challenge experiments, in which animals vaccinated with S107 cells mixed with CL7.1/mCD40L fibroblasts were subsequently challenged with live MPC-11 cells.Vaccination with apoptotic/necrotic tumor cells Apoptotic tumor cells were also used as a source of tumor-associated antigens for immunization. For all plasmacytoma cell lines, apoptotic tumor cells were obtained after exposure to UV irradiation for 60 seconds and culture overnight in serum-free conditions. Apoptotic cells obtained from a starting culture of 2 × 106 tumor cells were washed in PBS, combined with either CL7.1/mCD40L fibroblasts or CL7.1/neo fibroblasts (ratio, 2:1), resuspended in 200 µL final volume, and injected SC in the right leg on day 1. At 2 weeks later, these animals received a second, identical SC injection in the opposite leg and, on day 28, were challenged on the back with live tumor cells as described above. Tumor growth was followed by measurement of the 2 maximum diameters of the tumor at the site of challenge. Mice were killed when the tumor size was larger than 10% of the animal's body weight or had induced distress or ulceration.Culture experiments Irradiated CL7.1 fibroblasts expressing either the mCD40L or the neo genes were resuspended in medium and seeded in 6-well plates at 2 × 105 cells per well. After overnight incubation, exponentially growing plasmacytoma cells (MPC-11, S107, or X-24) were added at different ratios in 5 mL final volume in the presence of the appropriate medium. Cells were recovered for phenotypic analysis after 24, 48, and 72 hours of culture at 37°C in 5%CO2. For the proliferation assay, CL7.1/neo and CL7.1/mCD40L fibroblasts were-irradiated (1000 rad) and placed
in a 96-well plate (1 × 105 cells per well). MPC-11,
S107, or X-24 cells, resuspended in the appropriate medium, were added
to the wells at different ratios in a total volume of 200 µL. After
24, 48, and 72 hours of culture in a humidified atmosphere containing
5% CO2 at 37°C, cells were pulsed with 0.5 µCi (0.018 MBq) methyl-3H-thymidine (Amersham, Braunschweing,
Germany) and cultured for additional 15 hours. The cells were harvested
onto filters and dried, and thymidine uptake was measured in a
 -scintillation counter (TrisCarb 2500 TR; Packard BioScience,
Downers Grove, IL). Three replicates were used for each condition.
Immunophenotype and detection of apoptotic cells Next, 50 000 tumor cells, fibroblasts, or mouse splenocytes were stained with phycoerythrin (PE)-conjugated or fluorescein isothiocyanate (FITC)-conjugated antibodies. The antimouse antibodies were as follows: CD3 (17A2), CD4 (L3T4), CD8a (Ly-2), CD80 (B7-1), CD86 (B7-2), CD154 (mCD40L, gp39), H-2Kd (anti-MHC class I), I-Ad/I-Ed (anti-MHC class II), and CD40 (clones 3/23 and HM40-3) (all from Pharmingen, San Diego, CA). Control samples labeled with an appropriate isotype-matched antibody (Pharmingen) were added to each experiment. After staining for 30 minutes at 4°C, cells were washed, fixed in paraformaldehyde 0.5%, and evaluated on a FACScan analyzer (Becton Dickinson, San Jose, CA) equipped with the filter set for triple fluorescence signals. Apoptosis was measured by cell staining with FITC-annexin-V (Pharmingen) antibody and propidium iodide, according to the manufacturer's instruction.Reverse transcription-polymerase chain reaction amplification Total RNA was extracted from plasmacytoma cells, mouse splenocytes, and fibroblasts by means of the Qiagen kit (Valencia, CA) according to the manufacturer's instructions. Then, 2 µg RNA were reverse transcribed to complementary DNA (cDNA) by means of the murine leukemia virus reverse transcriptase (GIBCO BRL). Polymerase chain reaction (PCR) amplification was performed in a final volume of 50 µL with the use of 2 µL cDNA, 0.25 mM deoxynucleoside 5'-triphosphate, 0.5 µM each primer, and 2 U AmpliTaq Gold Polymerase (Perkin-Elmer Cetus, Norwalk, CT) in PCR buffer. One cycle at 95°C for 6 minutes followed by 30 cycles, each consisting of 60 seconds at 94°C, 60 seconds at 56°C, and 90 seconds at 72°C, were performed in a Perkin-Elmer Cetus DNA Thermal Cycler. The primers for the murine CD40 amplification were 5'-ACGTGCAGTGACAAACAGTAC-3' (sense) and 5'-CACAGCTTGTCCAGGGATAAC-3' (antisense). The integrity of RNA extracted was evaluated in each experiment by amplification of the murine-actin. The reaction
products were electrophoretically separated through a 1.5% agarose gel
and stained with ethidium bromide. The expected products generated by
PCR were 429 base pairs (bp) and 462 bp for CD40 and -actin,
respectively. To exclude contamination, appropriate negative controls
were added to each experiment.
Statistical analysis All data are presented as mean ± 1 SD. Student t test was used to determine the statistical significance of differences between samples, and P < .05 was accepted as indicating significance. Tumor-free survival was analyzed by Kaplan-Meier analysis, and the statistical significance of observed differences was assessed by log-rank testing.
Transgenic expression of CD40L reduces the in vivo tumor growth of CD40+ plasmacytoma cell lines As shown in Figure 1A, the murine plasmacytoma cell lines MPC-11 and S107 expressed CD40 antigen on their surface, as detected by staining with anti-CD40 mAb (clone HM40-3). Addition of CD40L-expressing fibroblasts did not significantly modulate the surface expression of CD40 on either cell line (Figure 1B), and the expression of CD40 on MPC-11 and S107 cells growing in vitro was essentially identical to the expression detected on cells growing in vivo in BALB/CBYJ mice (36 ± 14 percent of positive cells and 32 ± 11 percent of positive cells, respectively, versus 40 ± 3 percent of positive cells and 27 ± 15 percent of positive cells, respectively).
To evaluate the effects of transgenic CD40L expression on plasmacytoma
cell growth in vivo, mice were inoculated SC with sublethally irradiated MPC-11 or S107 cells mixed with irradiated CL7.1/mCD40L or
CL7.1/neo fibroblasts (ratio, 2:1). Tumor growth was followed at the injection site. For both MPC-11 and S107 cell lines, transgenic expression of CD40L significantly reduced tumor growth compared with
control animals receiving tumor cells mixed with CL7.1/neo fibroblasts
(P = .008 and P < .001, respectively)
(Figure 2A-B). At 6 weeks after tumor
injection, all control animals (8 of 8) receiving sublethally
irradiated MPC-11 or S107 cells mixed with CL7.1/neo fibroblasts were
euthanized because of massive tumor growth. In contrast, only 2 of 8 animals injected with sublethally irradiated MPC-11 or S107 cells mixed
with CL7.1/mCD40L fibroblasts had measurable tumors (Table
1).
We next excluded the possibility that improved survival was due to the direct inhibitory effect of CD40L transgene expression on tumor cell growth, mediated through tumor CD40 stimulation. MPC-11 or S107 cells were cocultured in vitro with CL7.1/mCD40L fibroblasts or CL7.1/neo fibroblasts at different ratios. As shown in Figure 2, transgenic expression of CD40L did not significantly influence the proliferative activity of either MPC-11 (Figure 2C) or S107 (Figure 2D) cell lines. Transgenic expression of CD40L induces immune-mediated tumor protection To evaluate the role of T lymphocytes in reducing tumor growth in animals receiving tumor cells and CL7.1/mCD40L fibroblasts, we selectively depleted CD8+ or CD4+ T lymphocytes in vivo before injection of the tumor cells. The results shown in Table 2 demonstrate that both CD8+ and CD4+ T lymphocytes contributed to the protective response. All animals (5 of 5) receiving MPC-11 or S107 cells mixed with CL7.1/mCD40L fibroblasts and depleted of CD8+ T lymphocytes by IP injection of 2.43 mAb were killed within 6 weeks because of massive tumor growth (Table 2). Similarly, when CD4+ T lymphocytes were depleted by IP injection of Gk1.5 mAb, the protective effect of CD40L transgenic expression against tumor growth was abrogated (Table 2).
The generation of CD4/CD8-mediated immunity is protective against subsequent tumor challenge Mice receiving 2 consecutive injections of sublethally irradiated tumor cells mixed with CL7.1/mCD40L fibroblasts were challenged with live tumor cells (5 × 105 cells) 2 weeks after the second injection. As shown in Figure 3, mice immunized in the presence of transgenically expressed CD40L had significantly increased tumor-free survival compared with controls receiving either MPC-11 (Figure 3A) or S107 (Figure 3B) cell lines (P = .0025 and P = .0003). All control mice (8 of 8) injected SC with live MPC-11 tumor cells developed fast-growing tumors and were killed within 3 weeks of the tumor injection, while only 3 of 8 mice vaccinated with CD40L had discernible tumor after challenge. Similarly, only 1 of 8 mice vaccinated with the S107 cell line and CL7.1/mCD40L fibroblasts developed tumor after challenge. Immunization with CD40L fibroblasts alone had no protective effect (data not shown). Cross-challenge experiments were also performed to analyze the specificity of the antitumor response. Of 13 mice injected twice with sublethally irradiated S107 cell line mixed with CL7.1/CD40L fibroblasts, 10 developed a discernable tumor when challenged with live MPC-11 cells; this was statistically significant compared with animals rechallenged with the S107 cell line (P = .009) (Figure 3C).
Tumor protection is not mediated by the acquisition of antigen-presenting function by CD40 activation of plasmacytoma cell lines CD40-stimulated lymphoma/CLL cells acquire improved antigen-presenting function.27,28 To evaluate whether the immune protection obtained against plasmacytoma cells following exposure to transgenic CD40L was also due to the acquisition of antigen-presenting function, we measured the expression of costimulatory molecules on plasmacytoma cells. Tumor cells and normal splenocytes were cocultured in vitro with fibroblasts expressing CD40L and analyzed daily for up-regulation of CD80 and CD86 costimulatory molecules and of MHC class II molecules. As shown in Figure 4 and Table 3, neither CD80/CD86 nor MHC class II molecules were up-regulated on either the MPC-11 or the S107 cell lines. In contrast, significant up-regulation of the CD80 molecule was observed in normal splenocytes cultured in the presence of CD40L-expressing fibroblasts, demonstrating the functionality of the transgenic molecule (Table 3).
Transgenic expression of CD40L mediates tumor protection in a
CD40  plasmacytoma cell
line. As shown in Figure 5, the X-24
plasmacytoma cell line is negative for CD40 messenger RNA (mRNA)
expression as assessed by reverse-transcription PCR analysis (Figure
5A) and for surface expression of the CD40 molecule as assessed by the
use of 2 different anti-CD40 mAbs (clones HM40-3 and 3/23) (Figure 5B).
The expression of CD40 on X-24 cells growing in vivo in BALB/CBYJ mice
was also undetectable (data not shown), and there was no measurable
up-regulation of CD40 from the addition of CD40L-expressing fibroblasts
(Figure 5C). As observed for the MPC-11 and S107 cells, transgenic
expression of CD40L by fibroblasts significantly reduced the local
growth of X-24 cells in BALB/CBYJ mice (Figure
6A) although transgenic expression of
CD40L did not significantly influence the proliferative activity in
vitro of X-24 cells compared with control cells cultured in the
presence of CL7.1/neo fibroblasts (data not shown). The local
inhibitory effect produced in vivo by transgenic CD40L expression was
immune mediated, since it induced significant protection when animals were challenged 2 weeks later with live tumor cells (Figure 6B). As
expected, no significant modulation of the costimulatory molecules (CD80 and CD86) was observed after coculture of CD40 X-24
cells with fibroblasts expressing CD40L (data not shown).
Immune-mediated tumor protection is determined predominantly by in vivo activation of professional APCs Because the protective effect induced by transgenic expression of CD40L was independent of the acquisition of antigen-presenting capability by plasmacytoma cells, we investigated whether this effect could be mediated by activation of professional APCs cross-primed with tumor-derived antigens. We injected animals with plasmacytoma cells previously exposed to UV radiation and cultured overnight in serum-free medium. Staining with annexin-V and propidium iodide and FACS analysis showed that more than 60% of cells were apoptotic and 25% necrotic, with fewer than 15% viable (negative for both annexin-V and propidium iodide). After mixing with CL7.1/mCD40L fibroblasts, this cell suspension was injected as previously described for viable tumor cells. Mice were challenged 2 weeks later with live tumor cells, and as shown in Figure 7, transgenic expression of CD40L produced the same systemic tumor protection observed in mice that had received viable tumor cells. Thus, 3 of 8 mice vaccinated with apoptotic MPC-11 cells and CL7.1/mCD40L fibroblasts developed tumor after challenge as compared with 8 of 8 controls (P = .0024) (Figure 7A). Similar levels of protection were observed when the S107 cell line (3 of 8 versus 8 of 8, P = .0003) (Figure 7B) or X-24 cell line (2 of 15 versus 7 of 8, P = .001) (Figure 7C) were substituted. Mice injected with the identical tumor cell suspensions mixed with CL7.1/neo fibroblast were not protected, because all mice developed tumor at the site of injection (data not shown).
In this report, we have shown that transgenic expression of CD40L in the proximity of MM cells can generate both local and systemic tumor protection. This effect is probably mediated by in vivo recruitment/activation of professional APCs able to take up antigens released by tumor cells, rather than by the acquisition of APC function by MM cells, and requires both CD4+ and CD8+ T lymphocytes. Vaccination with whole tumor cells represents an attractive strategy for human cancer immunotherapy.12 Although the substitution of defined tumor-associated antigens may ultimately be desirable, human tumors are extremely heterogeneous, making the identification of one or more "universal" tumor antigens extremely difficult for many tumors. In addition, many potential tumor-associated antigens have yet to be identified.32 Finally, it is becoming increasingly evident that the use of defined antigens may be able to recruit only a limited humoral or cellular immune response, one that is inadequate to produce sustained and effective antitumor activity in humans. Instead, it is necessary to use antigenic stimuli that recruit both CD8+ and CD4+ T lymphocytes that can each recognize their relevant class I and class II MHC-restricted tumor epitopes residing on a single or proximate APC.33 These critical cognate interactions between helper and cytotoxic T cells may be difficult to achieve with a single defined tumor antigen, which may lack peptide sequences capable of being presented by both class I and class II MHC molecules on the patient's APCs. Transgenic expression of CD40L is one means of enhancing the immune response to whole tumor cells, and the molecule may be provided on the tumor cell itself or, as here, by bystander cells in the vicinity of the injected tumor.21 CD40L is particularly attractive for the immunotherapy of B-cell tumors because these are often CD40+. It has been suggested that in this setting, CD40 activation by CD40L up-regulates the costimulatory molecules (CD80/B7.1 and CD86/B7.2) and class II MHC expression on the malignant cells, enhancing their inherent antigen-presenting capabilities.21,27,28 This in turn increases the possibility of generating specific cytotoxic T lymphocytes.34 Our results with MM show that transgenic expression of CD40L by syngeneic fibroblasts in the tumor microenvironment reduces the local tumor growth of plasmacytoma cell lines. This local effect is not dependent on any direct inhibitory effect of CD40L expression on the growth of either tumor.35-37 Instead, it is immune mediated, because vaccination inhibited not only local tumor growth, but also tumor growth from a subsequent tumor challenge at a distal site. Importantly, immune protection requires both CD4+ and CD8+ T lymphocytes, because depletion of either subset abolishes protection. These results indicate that transgenic CD40L expression has been associated with the processing and subsequent presentation of peptides derived from antigens released by tumor cells to both CD4+ and CD8+ effector cells. This combination of cellular responses may particularly favor the broad and sustained antitumor responses required for the treatment of human tumors. Transgenic CD40L had no effect on MHC or costimulatory molecule (CD80
and CD86) expression by CD40+ plasmacytoma cells,
indicating that the immune effects were unlikely to be mediated by an
increase in the ability of the tumor to function as an APC consequent
upon CD40 activation. CD40L also fails to enhance tumor IL-6 release in
both the MPC-11 and the S107 cell line, even when the transgenic CD40L
highly up-regulates this cytokine production when added to spleen cells
(data not shown). This contrasts with findings observed in other B-cell
malignancies, such as lymphoma/CLL,27,28 and suggests that
these murine MM cells could be less responsive to CD40-CD40L signaling.
This deficit may be a general phenomenon in MM since CD40 activation by
transgenic CD40L also fails to up-regulate B7 molecules in human MM
cell lines35,36 and primary MM cells.38
Although a normal capacity to up-regulate alternative costimulatory
molecules on plasmacytoma cells cannot be ruled out, MM cells appear to
have a defect in the CD40 transduction pathway. Cross-linking of CD40
on normal B lymphocytes induces the activation of several pathways,
including Rel/nuclear factor- In conclusion, we have shown that transgenic CD40L expression induces a
protective response against myeloma. Both CD4+ and
CD8+ tumor-specific T cells are induced, a combination
likely to be desirable for the immunotherapy of human
malignancy.12,32 Moreover, because our results show that
the interaction with CD40 antigen on tumor cells is neither universal
nor required for the immune enhancement mediated by transgenic CD40L,
the molecule may be of therapeutic value in both CD40+ and
CD40
We thank Susan Smith and Tatiana Gotsolova for excellent technical assistance. We also thank Gloria Levin for help in preparing this manuscript.
Submitted February 6, 2001; accepted February 12, 2002.
Supported by National Institutes of Health Grants RO1 CA75014 and RO1
CA78792; G.D. was supported by a grant from Associazione Paolo
Belli
G.D. and B.S. equally contributed to this work.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: Malcolm K. Brenner, Center for Cell and Gene Therapy, Baylor College of Medicine, 6621 Fannin St, MC 3-3320, Houston, TX 77030; e-mail: mbrenner{at}bcm.tmc.edu.
1.
Attal M, Harousseau JL, Stoppa AM, et al.
A prospective randomized trial of autologous bone marrow transplantation and chemotherapy in multiple myeloma.
N Engl J Med.
1996;335:91-97
2.
Lenhoff S, Hjorth M, Holmberg E, et al.
Impact on survival of high-dose therapy with autologous stem cell support in patients younger than 60 years with newly diagnosed multiple myeloma: a population-based study. Nordic Myeloma Study Group.
Blood.
2000;95:7-11
3.
Sirisinha S, Eisen HN.
Autoimmune-like antibodies to the ligand-binding sites of myeloma proteins.
Proc Natl Acad Sci U S A.
1971;68:3130-3135
4.
Dianzani U, Pileri A, Boccadoro M, et al.
Activated idiotype-reactive cells in suppressor/cytotoxic subpopulations of monoclonal gammopathies: correlation with diagnosis and disease status.
Blood.
1988;72:1064-1068
5.
Yi Q, Osterborg A, Bergenbrant S, Mellstedt H, Holm G, Lefvert AK.
Idiotype-reactive T-cell subsets and tumor load in monoclonal gammopathies.
Blood.
1995;86:3043-3049
6.
Treon SP, Mollick JA, Urashima M, et al.
Muc-1 core protein is expressed on multiple myeloma cells and is induced by dexamethasone.
Blood.
1999;93:1287-1298
7.
van Baren N, Brasseur F, Godelaine D, et al.
Genes encoding tumor-specific antigens are expressed in human myeloma cells.
Blood.
1999;94:1156-1164
8.
Massaia M, Borrione P, Battaglio S, et al.
Idiotype vaccination in human myeloma: generation of tumor-specific immune responses after high-dose chemotherapy.
Blood.
1999;94:673-683 9. King CA, Spellerberg MB, Zhu D, et al. DNA vaccines with single-chain Fv fused to fragment C of tetanus toxin induce protective immunity against lymphoma and myeloma. Nat Med. 1998;4:1281-1286[CrossRef][Medline] [Order article via Infotrieve].
10.
Reichardt VL, Okada CY, Liso A, et al.
Idiotype vaccination using dendritic cells after autologous peripheral blood stem cell transplantation for multiple myeloma: a feasibility study.
Blood.
1999;93:2411-2419
11.
Li Y, Bendandi M, Deng Y, et al.
Tumor-specific recognition of human myeloma cells by idiotype-induced CD8(+) T cells.
Blood.
2000;96:2828-2833 12. Ostrand-Rosenberg S, Pulaski BA, Clements VK, Qi L, Pipeling MR, Hanyok LA. Cell-based vaccines for the stimulation of immunity to metastatic cancers. Immunol Rev. 1999;170:101-114[CrossRef][Medline] [Order article via Infotrieve].
13.
Simons JW, Jaffee EM, Weber CE, et al.
Bioactivity of autologous irradiated renal cell carcinoma vaccines generated by ex vivo granulocyte-macrophage colony-stimulating factor gene transfer.
Cancer Res.
1997;57:1537-1546
14.
Soiffer R, Lynch T, Mihm M, et al.
Vaccination with irradiated autologous melanoma cells engineered to secrete human granulocyte-macrophage colony-stimulating factor generates potent antitumor immunity in patients with metastatic melanoma.
Proc Natl Acad Sci U S A.
1998;95:13141-13146
15.
Bowman L, Grossmann M, Rill D, et al.
IL-2 adenovector-transduced autologous tumor cells induce antitumor immune responses in patients with neuroblastoma.
Blood.
1998;92:1941-1949
16.
Huang AY, Golumbek P, Ahmadzadeh M, Jaffee E, Pardoll D, Levitsky H.
Role of bone marrow-derived cells in presenting MHC class I-restricted tumor antigens.
Science.
1994;264:961-965
17.
Noelle RJ, Roy M, Shepherd DM, Stamenkovic I, Ledbetter JA, Aruffo A.
A 39-kDa protein on activated helper T cells binds CD40 and transduces the signal for cognate activation of B cells.
Proc Natl Acad Sci U S A.
1992;89:6550-6554
18.
Caux C, Massicrier C, Vanbervliet B, et al.
Activation of human dendritic cells through CD40 cross-linking.
J Exp Med.
1994;180:1263-1272 19. Ridge JP, Di Rosa F, Matzinger P. A conditioned dendritic cell can be a temporal bridge between a CD4+ T-helper and T-killer cell. Nature. 1998;393:474-478[CrossRef][Medline] [Order article via Infotrieve]. 20. Schoenberger SP, Toes RE, van der Voort EI, Offringa R, Melief CJ. T-cell help for cytotoxic T lymphocytes is mediated by CD40-CD40L interactions. Nature. 1998;393:480-483[CrossRef][Medline] [Order article via Infotrieve].
21.
Dilloo D, Brown M, Roskrow M, et al.
CD40 ligand induces an antileukemia immune response in vivo.
Blood.
1997;90:1927-1933 22. Grossmann ME, Brown MP, Brenner MK. Antitumor responses induced by transgenic expression of CD40L. Hum Gen Ther. 1997;9:1935-1943. 23. Couderc B, Zitvogel L, Douin-Echinard V, et al. Enhancement of antitumor immunity by expression of CD70 (CD27 ligand) or CD154 (CD40 ligand) costimulatory molecules in tumor cells. Cancer Gene Ther. 1998;5:163-175[Medline] [Order article via Infotrieve].
24.
Townsend SE, Allison JP.
Tumor rejection after direct costimulation of CD8+ T cells by B-7 transfected melanoma cells.
Science.
1993;259:368-370 25. Chen L, Ashe S, Brady WA, et al. Costimulation of antitumor immunity by the B7 counterreceptor for the T lymphocyte molecules CD28 and |
plasmacytoma cells
Top
Abstract
Introduction
B factors, a family of transcription
factors involved in the control of several genes crucial for lymphoid cell functions.
for example, the
failure of line S107 to bind to a second anti-CD40 mAb clone, 3/23
(data not shown). But although the molecular mechanism(s) of the absent up-regulation of CD80 and CD86 costimulatory molecules after CD40-CD40L engagement in plasmacytoma may be complex and varied, the absence of
these effects and the ability to use transgenic CD40L to obtain protection from a CD40